Identification of proSAAS homologs in lower vertebrates: conservation of hydrophobic helices and convertase-inhibiting sequences.
The prohormone convertases (PCs) 1/3 and 2 accomplish the major proteolytic cleavage events in neuroendocrine tissues; each of these convertases has a small associated binding protein that inhibits convertase action in the secretory pathway. The proSAAS protein binds to PC1/3, whereas the 7B2 protein binds to PC2. However, both convertase-binding proteins are more widely expressed than their cognate enzymes, suggesting that they may perform other functions as well. All known mammalian proSAASs are over 85% conserved; thus, identifying functionally important segments has been impossible. Here, we report the first identification of nonmammalian proSAAS molecules, from Xenopus and zebrafish (Danio rerio). Although these two proteins show an overall amino acid sequence identity of only 29 and 30% with mouse proSAAS, two 14-16 residue hydrophobic segments (predicted to form alpha-helices) and two, nine through 11 residue sequences containing basic convertase cleavage sites are highly conserved; therefore, these sequences may be of functional importance. Confidence that these nonmammalian molecules represent authentic proSAAS is supported by the finding that both inhibit mouse PC1/3 with nanomolar inhibition constants; human furin was not inhibited. In vitro, the two proteins were cleaved by PC2 and furin to three or more peptide products. Both zebrafish and Xenopus proSAAS exhibited neural and endocrine distributions, as assessed by in situ and PCR experiments, respectively. In summary, the identification of proSAAS molecules in lower vertebrates provides clues as to functional regions within this widely expressed neuroendocrine protein.
Kudo
H
H
Department of Anatomy and Neurobiology, University of Maryland-Baltimore, Baltimore, Maryland 21201, USA.
Liu
J
J
Jansen
E J R
EJ
Ozawa
A
A
Panula
P
P
Martens
G J M
GJ
Lindberg
I
I
D000595:Amino Acid Sequence; D000818:Animals; D017124:Conserved Sequence; D004625:Embryo, Nonmammalian; D004791:Enzyme Inhibitors / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D008969:Molecular Sequence Data; D055099:Neuroendocrine Cells / Q000378:metabolism; D009474:Neurons / Q000378:metabolism; D009479:Neuropeptides / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D010802:Phylogeny; D043484:Proprotein Convertases / Q000037:antagonists & inhibitors*; D017433:Protein Structure, Secondary / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D014714:Vertebrates / Q000235:genetics*; D014982:Xenopus laevis / Q000235:genetics; D015027:Zebrafish / Q000235:genetics
D004791:Enzyme Inhibitors; D009479:Neuropeptides; C403532:PCSK1N protein, human; D043484:Proprotein Convertases